Structural insight into nucleosome transcription by RNA polymerase II with elongation factors

Ehara, Haruhiko; Kujirai, Tomoya; Fujino, Y.; Shirouzu, Mikako; Kurumizaka, Hitoshi; Sekine, S.

doi:10.1126/science.aav8912

Cited by 135 publications

(154 citation statements)

References 40 publications

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“…Very recently, cryo-EM structures of Pol II-nucleosome complexes have provided snapshots of Pol II paused at major histone-DNA contacts and suggested sites of interaction with other factors 31–33 . Missing from these structures is information about the dynamics of barrier crossing by the enzyme: what are the time windows available for in-trans interactions with these discrete sites, how are these related to the local energetic magnitude of the barrier, and how are they modulated by epigenetic modifications and histone variants.…”

Section: Discussionmentioning

confidence: 99%

High-resolution and High-accuracy Topographic and Transcriptional Maps of the Nucleosome Barrier

Chen

Gabizon

Bustamante

et al. 2019

Preprint

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Nucleosomes represent mechanical and energetic barriers that RNA Polymerase II (Pol II) must overcome during transcription. A high-resolution description of the barrier topography, its modulation by epigenetic modifications, and their effects on Pol II nucleosome crossing dynamics, is still missing. Here, we obtain topographic and transcriptional (Pol II residence time) maps of canonical, H2A.Z, and monoubiquitinated H2B (uH2B) nucleosomes at near base-pair resolution and accuracy. Pol II crossing dynamics are complex, displaying pauses at specific loci, backtracking, and nucleosome hopping between wrapped states. While H2A.Z widens the barrier, uH2B heightens it, and both modifications greatly lengthen Pol II crossing time. Using the dwell times of Pol II at each nucleosomal position we extract the energetics of the barrier. The orthogonal barrier modifications of H2A.Z and uH2B, and their effects on Pol II dynamics rationalize their observed enrichment in +1 nucleosomes and suggest a mechanism for selective control of gene expression. Highlights A single-molecule unzipping assay mimics DNA unwinding by Pol II and maps the topography of human canonical, H2A.Z and uH2B nucleosome barriers at high resolution Real-time dynamics and full molecular trajectories of Pol II crossing the nucleosomal barrier reveal the transcriptional landscape of the barrier at high accuracy H2A.Z enhances the width and uH2B the height of the barrier A unified mechanical model links position-dependent dwell times of Pol II on the nucleosome with energetics of the barrier

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Section: Discussionmentioning

confidence: 99%

High-resolution and High-accuracy Topographic and Transcriptional Maps of the Nucleosome Barrier

Chen

Gabizon

Bustamante

et al. 2019

Preprint

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“…For transcription, therefore, the DNA wrapped around the octamer must be unwrapped. This is accomplished by RNA polymerase II, which unwraps the nucleosomal DNA in stepwise fashion during the transcription elongation process (2,3).…”

Section: Introductionmentioning

confidence: 99%

Structural Studies of Overlapping Dinucleosomes in Solution

Matsumoto

Sugiyama

et al. 2020

Biophysical Journal

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An overlapping dinucleosome (OLDN) is a structure composed of one hexasome and one octasome and appears to be formed through nucleosome collision promoted by nucleosome remodeling factor(s). In this study, the solution structure of the OLDN was investigated through the integration of small-angle x-ray and neutron scattering (SAXS and SANS, respectively), computer modeling, and molecular dynamics simulations. Starting from the crystal structure, we generated a conformational ensemble based on normal mode analysis and searched for the conformations that reproduced the SAXS and SANS scattering curves well. We found that inclusion of histone tails, which are not observed in the crystal structure, greatly improved model quality. The obtained structural models suggest that OLDNs adopt a variety of conformations stabilized by histone tails situated at the interface between the hexasome and octasome, simultaneously binding to both the hexasomal and octasomal DNA. In addition, our models define a possible direction for the conformational changes or dynamics, which may provide important information that furthers our understanding of the role of chromatin dynamics in gene regulation.

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“…NCP unwrapping has been visualised with cryo-EM, for example in the context of the hexasome, which is an NCP with partially unpeeled DNA, due to the loss of one H2A/H2B dimer 27 . Spectacular views of progressively unwrapped NCPs have been obtained for transcribing RNA polymerase II captured during NCP passage 43,44 . Moreover, cryo-EM provided the first glimpses of ATP-dependent NCP translocation through a mechanism involving DNA looping and sliding along the histone octamer 22,38,45–51 .…”

Section: Discussionmentioning

confidence: 99%

Retroviral integration into nucleosomes through DNA looping and sliding along the histone octamer

Wilson

Renault

Maskell

et al. 2019

Preprint

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24Retroviral integrase can efficiently utilise nucleosomes for insertion of the reverse-25 transcribed viral DNA. In face of the structural constraints imposed by the nucleosomal 26 structure, integrase gains access to the scissile phosphodiester bonds by lifting DNA off the 27 histone octamer at the site of integration. To clarify the mechanism of DNA looping by 28 integrase, we determined a 3.9 Å resolution structure of the prototype foamy virus intasome 29 engaged with a nucleosome core particle. The structural data along with complementary 30 single-molecule Förster resonance energy transfer measurements reveal twisting and sliding 31 of the nucleosomal DNA arm proximal to the integration site. Sliding the nucleosomal DNA 32 by approximately two base pairs along the histone octamer accommodates the necessary 33 DNA lifting from the histone H2A-H2B subunits to allow engagement with the intasome. 34Thus, retroviral integration into nucleosomes involves the looping-and-sliding mechanism for 35 nucleosomal DNA repositioning, bearing unexpected similarities with chromatin remodelers. 36 37 38 42 function, a multimer of IN assembles on viral DNA (vDNA) ends forming a highly stable 43 nucleoprotein complex, known as the intasome 2-4 . In its first catalytic step, IN resects 3' ends 44 of the vDNA downstream of the invariant CA dinucleotides (3'-processing reaction). It then 45 utilises the freshly released 3'-hydroxyl groups as nucleophiles to attack a pair of 46 phosphodiester bonds on opposing strands of chromosomal DNA, cleaving host DNA and 47 simultaneously joining it to 3' vDNA ends (strand transfer reaction) 5,6 .48 Many important questions pertaining to the nature of the host-virus transactions on 49 chromatin remain unanswered. In particular, it is unclear what role chromatin structure plays 50 in the integration process. Strikingly, although only a fraction of the nucleosomal DNA 51 surface is exposed within the nucleosome core particle (NCP) 7-9 , nucleosomal DNA packing 52 does not impede and rather stimulates integration 10-15 . Because retroviral INs have long 53 been known to prefer bent or distorted DNA targets, DNA bending as it wraps around the 54 histone octamer was thought to facilitate integration into NCPs 12,13 . However, recent 55 structural data revealed that retroviral intasomes require target DNA to adopt a considerably 56 sharper deformation than the smooth bend observed on NCPs 15-19 .57 Intasome structures from several retroviral genera have been determined by X-ray 58 crystallography and cryo-EM 4,17-20 . Despite considerable variability, all intasomes were 59 found to contain the structurally conserved intasomal core assembly minimally comprising 60 four IN subunits synapsing a pair of vDNA ends. Depending on the retroviral species, the 61 core assembly can be decorated by a number of additional IN subunits. The nucleoprotein 62 complex from the prototype foamy virus (PFV) contains only a tetramer of IN, making this 63 well-characterised intasome an ideal model to study the basic mech...

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Structural insight into nucleosome transcription by RNA polymerase II with elongation factors

Cited by 135 publications

References 40 publications

High-resolution and High-accuracy Topographic and Transcriptional Maps of the Nucleosome Barrier

High-resolution and High-accuracy Topographic and Transcriptional Maps of the Nucleosome Barrier

Structural Studies of Overlapping Dinucleosomes in Solution

Retroviral integration into nucleosomes through DNA looping and sliding along the histone octamer

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